Effectiveness of GeneXpert® in the diagnosis of tuberculosis in people living with HIV/AIDS

ABSTRACT OBJECTIVE To identify and synthesize the scientific knowledge produced regarding the effectiveness of the GeneXpert test in the diagnosis of pulmonary tuberculosis (TB) in people living with HIV/AIDS. METHODS Integrative literature review, which was searched on Embase, Scopus, PubMed, Cinahl, Academic Search Premier, Socindex, and LILACS platforms, in December 2019. The studies surveyed went through two stages of selection: reading of titles and abstracts by two reviewers independently; using the Rayyan platform and reading. Nineteen primary studies in English, Portuguese, and Spanish that answered the study’s guiding question were included: How effective is the GeneXpert test in the diagnosis of pulmonary TB in people living with HIV/AIDS? RESULTS The use of GeneXpert substantially increased the detection of TB cases among the population co-infected with HIV/AIDS, with sensitivity ranging from 68% to 100%, superior to sputum smear microscopy. Specificity ranged from 91.7% to 100%; the positive predictive value from 79.2% to 96.1%; and the negative predictive value from 84.6% to 99.3%. These values were considered similar to sputum smear microscopy by most studies. We also compared these results with different ways of performing culture and other molecular tests, being considered inferior only to the Xpert Ultra. CONCLUSION It is possible to affirm that places with a high incidence of HIV/AIDS would benefit from the implementation of the GeneXpert test, entailing effectiveness in diagnosing pulmonary TB in this population when compared to sputum smear microscopy, a widely used test for detection of cases.


INTRODUCTION
Since the 1980s, infection by the human immunodeficiency virus (HIV) has reinforced the concern with tuberculosis (TB) 1  In the 1990s, TB control strategies were based on passive detection of the disease through sputum smear microscopy in patients with chronic cough. However, the clinical presentation of pulmonary TB among people living with HIV/AIDS is nonspecific in advanced stages, with less frequent coughing and negative sputum smears 3 .
Although this microbiological diagnostic technique offers advantages in terms of costs and simplicity, specificity and sensitivity are still considered precarious, especially among people living with HIV/AIDS 4,5 , given the scarce production of sputum and the decrease in the bacillary load 6 . In addition to microbiological results, the diagnosis of TB must be supported by the clinical presentation 7 and radiographic findings 8 .
Another microbiological test used for the diagnosis of TB, considered the gold standard for laboratory confirmation of the disease, is the sputum culture. This test has high sensitivity in detecting Mycobacterium tuberculosis (MTB) and has been recommended for people with HIV/AIDS 9 . However, performing such examination requires resources, qualified technical skills, and time to obtain results, which delays the diagnosis and increases the risk of disease transmission 10,11 .
So the diagnosis of TB in people living with HIV/AIDS was often based on the clinical presentation, considering the lack of microbiological proof of infections 12 . This led to the expectation of developing new diagnostic means for use by health services, among which the rapid molecular tests (TRM) 13 , such as the GeneXpert ® MTB/RIF (GeneXpert). This is a nucleic acid amplification-based test, based on polymerase chain reaction (PCR) whereby it detects MTB and its resistance to rifampicin in a sputum sample within a period of two hours 14 .
This test was approved in 2010 by the WHO, which recommended its use for the initial diagnosis of TB and suspected cases of multidrug-resistant tuberculosis (MDR-TB) in sputum samples 15 , and expanded this recommendation in 2013, also for the diagnosis of TB in non-respiratory samples, that is, for extrapulmonary TB [13][14][15][16] . In 2014, as a result of these recommendations, 3,269 GeneXpert devices were made available to public sectors in 108 of the 145 countries eligible to obtain the equipment 7 .
The descriptors in bold, mentioned in Box 1, are part of the controlled vocabulary consulted in the Health Sciences Descriptors (DECS), whereby it was also possible to identify the corresponding terms in Spanish and English. For terms in English, we also consulted the Medical Subject Headings (MeSH). Finally, the free vocabulary used in the writing of publications was also sought, searching for synonyms in DECS and MeSH and databases selected for the study.
We carried bibliographic survey out in December 2019 using the vocabulary found (Box 2) and the Boolean operators OR (among words with the same meaning) and AND (among groups of words with the same meaning).
The bibliographic survey was executed in December 2019, in the following databases: Excerpta Medica dataBASE (EMBASE), SciVerse Scopus (SCOPUS) on behalf of Elsevier, Public/Medline or Publisher Medlin (PubMed), Cumulative Index to Nursing and Allied Health Literature (CINAHL), Academic Search Premier (ASP), Bibliographic Database for Sociology Research (SocINDEX) and Literatura Latino-Americana e do Caribe em Ciências da Saúde (LILACS). For the LILACS search, we used the vocabulary in the three mentioned languages; in other databases, the English vocabulary was used. It is important to emphasize that we did not use publication year nor language limits in the bibliographic survey.
which identified and allowed the exclusion of 597 duplicate publications. Two independent reviewers judged the eligibility of the remaining publications by reading their titles and abstracts. The following criteria guided the inclusion of 19 studies in this review: primary studies; having people with TB-HIV co-infection as the study population; address the diagnosis of pulmonary TB; and studies that answered the guiding questions of this review. We also highlight that all studies considered in this review were only based on the GeneXpert technology produced by the company Cepheid. Articles that studied cost-effectiveness, abstracts published in annals, and articles that did not compare the GeneXpert test with other diagnostic tests were excluded (Figure).
Data from the articles included in this review were extracted using a specific instrument based on Ursi (2005) 21 , allowing the narrative synthesis of its results. Finally, the methodological quality of the articles was assessed using the "Checklist for Studies on the Accuracy of Diagnostic Tests", proposed by The Joanna Briggs Institute (JBI) (https://joannabriggs.org/critical-appraisal-tools). It includes ten evaluation items within dichotomous answers (yes/no), considering "yes" a positive answer. Two checklist items were not applicable to the selected studies, so we sorted them out during evaluations. being non-resistant to rifampicin, and 03 (2.00%) resistant to rifampicin. The percentage of positivity for sputum smear microscopy was 11.8% and for LJ culture was 21.7%. In comparison, researchers observed that the sensitivity, specificity, and agreement were above 90%, with no statistically significant differences between them. the American Continent 37,39 , one (5.3%), from Asia 24 and one (5.3%) from a multicenter study 35 that involves countries from Africa, America, Europe, and Asia. , one (5.3%) Light Cycler Mycobacterium Detection (LCTB) 6 and one (5.3%) MTBDRplus (Hain Lifescience) 6 . Three articles 30,32,33 evaluated effectiveness of the test according to the results of CD4+ T lymphocyte counts and one article 33 evaluated this effectiveness according to the time of sputum collection.

From
As for the methodological quality of the studies, five studies 23,25,26,37,38 included all the essential items considered by The Joanna Briggs Institute. Only one (5.3%) 27 article presented sample size calculation.

DISCUSSION
The epidemic and the expansion of HIV/AIDS infection impacted the incidence of TB 40 , whose early detection followed by timely treatment represent the key to its control, so that laboratory diagnosis represents a challenge for health services 39 . The population living with TB/HIV co-infection has some characteristics that limit the use of diagnostic tools, as it can more easily present a negative smear, advanced immunosuppression that provides delayed diagnosis, the subclinical manifestation of signs and symptoms characteristic of TB 41 .
In TB/HIV endemic regions with limited resources, sputum microscopy is usually the only method available for diagnosis 4,32,34,42 . The use of only this technique to screen for TB in the population co-infected with HIV restricts its potential diagnosis because approximately 10,000 organisms per milliliter are needed in the sputum to perform the test, an uncommon amount among the population living with HIV/AIDS 34 .
Due to these particularities, the relevance of sputum smear microscopy for the diagnosis of TB where GeneXpert is available is questioned, although evidence found shows that the concomitant use of both tests can result in an increase in the identification of cases not detected by both an exam and on the other 28,30 , mainly in people with T CD4+ lymphocyte count < 200 cells/µl 30 .
Other studies do not address the simultaneous performance of both tests but point out that the use of GeneXpert substantially increased the detection of TB cases among the population co-infected with HIV/AIDS 24 The WHO recommendation to use GeneXpert, instead of sputum smear microscopy as the initial test of choice in HIV-infected individuals susceptible to TB 6 , corroborates the findings of studies that have shown that GeneXpert's sensitivity in diagnosing TB in people living with HIV ranged from 68% 31 to 100% 34 (average 81.1%), being greater than the sensitivity of sputum smear microscopy 6,23,[25][26][27][30][31][32]34,37 , which ranged from 23.0% 26 to 66,7% 37 (average 43.3%). Given the above, the implementation of GeneXpert, a fast and accurate test in different clinical contexts, has led to a significant increase in the diagnosis of pulmonary TB, both when used as a first-choice test and as a complement 45 .
We highlight that sensitivity is calculated based on the sputum culture, which is considered the gold standard test for the diagnosis of TB. Although this is a reference test, some studies compare it with GeneXpert, pointing to lesser sensitivity of the latter in relation to culture using the LJ and BACTEC MGIT 960 methods 31 , while another study shows greater 23 sensitivity and other equal performance in relation to the LJ method 32 . Two studies brought other diagnostic methods that were also compared to GeneXpert: one indicated similar sensitivity between it and 6 the LCTB test, and another indicated lower sensitivity when compared to XpertUltra 35 . Thus, we highlight the importance of using GeneXpert as the test of choice for the diagnosis of pulmonary TB in adults living with HIV, because it has similar and superior effectiveness compared to culture and even to other molecular methods, such as LCTB. However, it is worth point out the importance of further studies in relation to the comparison of the test with culture, because divergences came up in relation to the results regarding such effectiveness. The indication of Xpert Ultra stands out as a method of choice to replace all available tests, including GeneXpert, because only one study was found in this review that compared the two tests 46 .
It should also be noted that the sensitivity of the GeneXpert test in people living with HIV was greater than in HIV-negative people 29 , in contrast to another study that stated that the sensitivity in people living with HIV was lower 30 . The increased sensitivity of the test in HIV-positive patients adds important advantages to its use in health services, such as the increase in the detection rate of microbiologically confirmed TB and the potential to institute a rapid treatment, aiming to reduce TB transmission 47 . Some individual characteristics, such as a negative sputum smear microscopy 26,29-31 and a CD4+ T lymphocyte count ≤ 200 cells/µl 32 , were mentioned as elements that reduce the sensitivity of the GeneXpert test. Although these results are controversial in relation to the study by Cavanaugh et al. (2016) 33 , which shows increased test sensitivity in people with lower CD4+ T lymphocyte counts, another study shows that the performance of GeneXpert in HIV-infected people with advanced immunosuppression has certain limitations 32 that may be caused by the lower concentration of mycobacteria in sputum and by the occult or subclinical presentation of the disease 30 .
The specificity of GeneXpert ranged from 91.7% 30 to 100% 34 (average 95.6%), and in comparison with sputum smear microscopy, some studies concluded that both exams showed similar performance in relation to the mentioned aspect 6,23,27,34,37 , and others that GeneXpert has less specificity 25,30 . Regarding other diagnostic tests, the study mentions that the specificity of the GeneXpert test is similar to that of culture 23 and LCTB 6 . The specificity of the test in people living with HIV/AIDS was also similar to people not infected by the virus 29 . Based on the above, it appears that the specificity of the GeneXpert test is comparable to other diagnostic tests.
The PPV, understood as the probability of having the disease when the test is positive, ranged from 79.2% to 96.1% (average 91%) for GeneXpert in people living with HIV/AIDS infection, is considered similar to sputum smear microscopy 6,25,27,34,37 and to LCTB 6 , as well as when the test is performed in people who are not living with the virus 30 , although one of the studies has pointed out that the PPV was lower in these people 29 . On the other hand, NPV, understood as the probability of not having the disease when the test is negative, ranged from 84.6% to 99.3% (average 92.6%) for GeneXpert in people living with HIV/AIDS, being considered similar 34,37 or greater 6,25,27 in relation to sputum smear microscopy and similar to LCTB 6 and in people who do not live with the virus 29 , although one of the studies has pointed out that the NPV is greater in these people 30 . Thus, as well as in the assessment of specificity, the results of the effectiveness of the GeneXpert test in terms of PPV and NVP showed evidence of being similar to the other comparison tests.
A study also presented the test's likelihood ratio for diagnosing TB in co-infected patients, calculated by dividing the probability of positive (positive likelihood) or negative (negative likelihood) results, in people with the disease, by the probability of the same result in people without the disease 49 . Regarding the positive likelihood ratio, there was no difference between GeneXpert and sputum smear microscopy 27 . However, the negative likelihood ratio of GeneXpert was lower and close to "zero" 27 , favoring it if compared to sputum smear microscopy.
The effectiveness of the test in identifying cases resistant to rifampicin needs to be further studied, as one study 25 showed that of seven identified cases, three were false positives; as well as the fact that other studies 6,22,24,32,37 have a very small population of resistant cases, limiting the conclusions that the test has similar effectiveness to sensitivity tests 22,24,32,37 and even to MDRTB plus 6 . So diagnosing resistance to rifampicin still remains a challenge for health services, considering it would reduce the average time to detect resistant cases to up to two days 32 , which would be an admirable performance when compared to almost 40 days of the test of sensitivity/conventional culture 25 . The immediate diagnosis of these cases would allow a substantial reduction in the risks of nosocomial transmission of the resistant bacillus and its specific and early treatment, which would result in a reduction in the severe forms of TB, as well as an improvement in the prognosis and a reduction in loss of clinical follow-up 25,39 .
The expansion of the GeneXpert test in diagnosing TB among HIV-infected individuals is essential, given its effectiveness and especially in countries with a high burden of coinfection, as this test would bring gains in case detection by replacing or complementing the sputum smear microscopy and would make the treatment time opportune 28 in relation to the culture, which has cost limitations and technical requirements 32,33 . We also emphasize that GeneXpert's diagnostic accuracy, based on 85% sensitivity and 97% specificity, would potentially save more than 400,000 lives per year 34 , even if some cases undergo empirical treatment, which would be reduced by locations with the implementation of testing 27,38 .
As for the methodological quality of the studies, five studies contemplated all the essential items considered by The Joanna Briggs Institute related to accuracy evaluation of tests 23,25,26,37,38 . Of the methodological limitations found in the studies, the small sample size stands out 22 ; sample losses above 20% 6,29,35 ; the lack of clarification regarding the sampling process 39 and the sampling process 22 ; the use of secondary data 32,36,39 ; doubts about performance 6,22,[27][28][29][30][32][33][34][35][36]39 and nonperformance 31 of blind studies in relation to the results of the tests performed; and the use of different reference tests for the study participants 36 or the use of sputum smear microscopy as a reference test 24,28,36 . Moreover, a possible population selection bias is identified in a study, within the GeneXpert test was only performed on samples that were positive for the culture 31 .
Despite the fact that the test is more costly than microscopy, the benefits of the time of diagnosis and indication of drug resistance proved to be greater, because they bring greater sensitivity and specificity. In addition, the test establishes the diagnosis in a significant proportion of smear-negative patients and has a cut-off consolidated for TB-MDR 30,45 . Based on this, GeneXpert can be used as a first-line diagnostic method 24 .
Some limitations should be noted. This review may not have included relevant studies that were not indexed in the searched databases, nor included the gray literature that could enable the identification of other topics/relevant points on the subject to be addressed.
It is possible to conclude that places with a high incidence of HIV/AIDS could benefit from the implementation of the GeneXpert test, because its effectiveness in diagnosing pulmonary TB in this population is expressive when compared to sputum smear microscopy, a test that has been used for a long time and widely used for case detection. When compared to other tests, there was controversy regarding the effectiveness of GeneXpert and the different types of cultures, similar to that of LCTB and lower than that of Xpert Ultra. In addition to increasing the detection of pulmonary TB cases among people living with HIV, the GeneXpert test can provide benefits such as faster results than those obtained by culture (the most sensitive method for confirming TB), culminating in timely detection and treatment, making GeneXpert a tool in the fight against TB. Along with the innovation in the use of GeneXpert, we stand for the support and strengthen of health services in the applicability of this technique in order to achieve the goals of sustainable development and the End TB Strategy.